ABSTRACT
BACKGROUND AND PURPOSE: Atherosclerosis is the primary pathological basis of cardiovascular disease. Ferroptosis is a regulated form of cell death, a process of lipid peroxidation driven by iron, which can initiate and promote atherosclerosis. STAT6 is a signal transducer that shows a potential role in regulating ferroptosis, but, the exact role in ferroptosis during atherogenesis remains unclear. The Traditional Chinese Medicine Maijitong granule (MJT) is used for treating cardiovascular disease and shows a potential inhibitory effect on ferroptosis. However, the antiatherogenic effect and the underlying mechanism remain unclear. In this study, we determined the role of STAT6 in ferroptosis during atherogenesis, investigated the antiatherogenic effect of MJT, and determined whether its antiatherogenic effect was dependent on the inhibition of ferroptosis. METHODS: 8-week-old male LDLR-/- mice were fed a high-fat diet (HFD) at 1st and 10th week, respectively, to assess the preventive and therapeutic effects of MJT on atherosclerosis and ferroptosis. Simultaneously, the anti-ferroptotic effects and mechanism of MJT were determined by evaluating the expression of genes responsible for lipid peroxidation and iron metabolism. Subsequently, we reanalyzed microarray data in the GSE28117 obtained from cells after STAT6 knockdown or overexpression and analyzed the correlation between STAT6 and ferroptosis. Finally, the STAT6-/- mice were fed HFD and injected with AAV-PCSK9 to validate the role of STAT6 in ferroptosis during atherogenesis and revealed the antiatherogenic and anti-ferroptotic effect of MJT. RESULTS: MJT attenuated atherosclerosis by reducing plaque lesion area and enhancing plaque stability in both preventive and therapeutic groups. MJT reduced inflammation via suppressing inflammatory cytokines and inhibited foam cell formation by lowering the LDL level and promoting ABCA1/G1-mediated lipid efflux. MJT ameliorated the ferroptosis by reducing lipid peroxidation and iron dysregulation during atherogenesis. Mechanistically, STAT6 negatively regulated ferroptosis by transcriptionally suppressing SOCS1/p53 and DMT1 pathways. MJT suppressed the DMT1 and SOCS1/p53 via stimulating STAT6 phosphorylation. In addition, STAT6 knockout exacerbated atherosclerosis and ferroptosis, which abolished the antiatherogenic and anti-ferroptotic effects of MJT. CONCLUSION: STAT6 acts as a negative regulator of ferroptosis and atherosclerosis via transcriptionally suppressing DMT1 and SOCS1 expression and MJT attenuates atherosclerosis and ferroptosis by activating the STAT6-mediated inhibition of DMT1 and SOCS1/p53 pathways, which indicated that STAT6 acts a novel promising therapeutic target to ameliorate atherosclerosis by inhibiting ferroptosis and MJT can serve as a new therapy for atherosclerosis treatment.
Subject(s)
Atherosclerosis , Cation Transport Proteins , Drugs, Chinese Herbal , Ferroptosis , STAT6 Transcription Factor , Suppressor of Cytokine Signaling 1 Protein , Animals , Ferroptosis/drug effects , Atherosclerosis/drug therapy , STAT6 Transcription Factor/metabolism , Male , Drugs, Chinese Herbal/pharmacology , Mice , Suppressor of Cytokine Signaling 1 Protein/metabolism , Tumor Suppressor Protein p53/metabolism , Signal Transduction/drug effects , Receptors, LDL/metabolism , Diet, High-Fat , Mice, Inbred C57BL , Mice, KnockoutABSTRACT
BACKGROUND: The rising incidence of metabolic diseases due to chronic inflammation in the adipose tissue has been attributed to factors such as high fat diet (HFD). Previous studies have demonstrated that the total saponins from Panax japonicus (TSPJ) can reduce HFD-induced adipocyte inflammation, but the underlying mechanism remains unclear. In this work, we explored the molecular mechanism by which TSPJ reduces inflammation response in adipocytes. METHODS: We first established C57BL/6 mouse and 3T3-L1 adipocyte models. Lentiviruses packaged with the plasmids were injected into mice through the tail vein or into adipocytes to generate the in vivo and in vitro models with miR155 knockdown and overexpression. The mice were fed with HFD to trigger inflammation and administered TSPJ (25 mg/kgâd and 75 mg/kgâd) by gavage. The adipocytes were treated with palmitic acid (PA) to trigger inflammation response, then treated with TSPJ (25 µg/ml and 50 µg/ml). Finally, the expression of miR155, inflammatory factors, SOCS1, and NFκB pathway-related proteins was explored. RESULTS: TSPJ significantly inhibited the expression of inflammation-related genes and the miR155 expression in adipocytes both in vitro and in vivo. The dual luciferase reporter gene assay revealed that miR155 mediated the downregulation of SOCS1. TSPJ significantly inhibited and upregulated the phosphorylation of the NFκB protein and the SOCS1 proteins, respectively. CONCLUSION: TSPJ inhibits miR155 to upregulate the SOCS1 expression, which subsequently inhibits the NFκB signaling pathway, thereby mitigating the inflammatory response in the adipocytes of HFD mice.
Subject(s)
MicroRNAs , Panax , Saponins , Mice , Animals , Saponins/metabolism , Mice, Inbred C57BL , Adipocytes/metabolism , Signal Transduction , NF-kappa B/metabolism , Inflammation/drug therapy , Inflammation/metabolism , Diet, High-Fat/adverse effects , 3T3-L1 Cells , Suppressor of Cytokine Signaling 1 Protein/metabolism , Suppressor of Cytokine Signaling 1 Protein/therapeutic use , MicroRNAs/genetics , MicroRNAs/metabolismABSTRACT
Wogonoside (WG) is a flavonoid chemical component extracted from Scutellaria baicalensis, which exerts therapeutic effects on liver diseases. Ferroptosis, a novel form of programmed cell death, regulates diverse physiological/pathological processes. In this study, we attempted to investigate a novel mechanism by which WG mitigates liver fibrosis by inducing ferroptosis in hepatic stellate cells (HSCs). A CCl4 -induced mouse liver fibrosis model and a rat HSC line were employed for in vivo and in vitro experiments, both treated with WG. Firstly, the levels of the fibrotic markers α-smooth muscle actin (α-SMA) and α1(I)collagen (COL1α1) were effectively decreased by WG in CCl4 -induced mice and HSC-T6 cells. Additionally, mitochondrial condensation and mitochondrial ridge breakage were observed in WG-treated HSC-T6 cells. Furthermore, ferroptotic events including depletion of SLC7A11, GPX4 and GSH, and accumulation of iron, ROS and MDA were discovered in WG-treated HSC-T6 cells. Intriguingly, these ferroptotic events did not appear in hepatocytes or macrophages. WG-elicited HSC ferroptosis and ECM reduction were dramatically abrogated by ferrostatin-1 (Fer-1), a ferroptosis inhibitor. Importantly, our results confirm that SOCS1/P53/SLC7A11 is a signaling pathway which promotes WG attenuation of liver fibrosis. On the contrary, WG mitigated liver fibrosis and inducted HSC-T6 cell ferroptosis were hindered by SOCS1 siRNA and pifithrin-α (PFT-α). These findings demonstrate that SOCS1/P53/SLC7A11-mediated HSC ferroptosis is associated with WG alleviating liver fibrosis, which provides a new clue for the treatment of liver fibrosis.
Subject(s)
Ferroptosis , Hepatic Stellate Cells , Animals , Mice , Rats , Liver , Liver Cirrhosis/drug therapy , Suppressor of Cytokine Signaling 1 Protein/metabolism , Suppressor of Cytokine Signaling 1 Protein/pharmacology , Suppressor of Cytokine Signaling 1 Protein/therapeutic use , Tumor Suppressor Protein p53/metabolismABSTRACT
Liver fibrosis is the consequence of chronic liver injury and is a major challenge to global health. However, successful therapy for liver fibrosis is still lacking. Sennoside A (SA), a commonly used clinical stimulant laxative, is reported to improve hepatic disease, but the underlying mechanisms remain largely elusive. Here, we show for the first time that SA enhanced suppressor of cytokine signaling 1 (SOCS1) expression in a DNA methyltransferase 1 (DNMT1)-dependent manner and thereby attenuated liver fibrosis. Consistently, SA inhibited the expression of the liver fibrogenesis markers α-smooth muscle actin (α-SMA) and type I collagen alpha-1 (Col1α1) and suppressed inflammatory responses in vivo and in vitro. Coculture experiments with macrophages/hepatic stellate cells (HSCs) revealed that SA suppressed HSC proliferation by downregulating proinflammatory cytokines in macrophages. Mechanically, SA promoted the aberrant expression of SOCS1 in liver fibrosis. However, blocking SOCS1 expression weakened the inhibitory effect of SA on HSC proliferation, indicating that SOCS1 may play an important role in mediating the antifibrotic effect of SA. Furthermore, SA inhibited DNMT1-mediated SOCS1 and reduced HSC proliferation by inhibiting inflammatory responses in carbon tetrachloride (CCl4) -induced liver fibrosis.
Subject(s)
Anti-Inflammatory Agents/therapeutic use , Chemical and Drug Induced Liver Injury/drug therapy , Liver Cirrhosis/drug therapy , Sennosides/therapeutic use , Suppressor of Cytokine Signaling 1 Protein/genetics , Animals , Anti-Inflammatory Agents/pharmacology , Carbon Tetrachloride , Cell Line , Cell Proliferation/drug effects , Chemical and Drug Induced Liver Injury/genetics , Chemical and Drug Induced Liver Injury/metabolism , DNA (Cytosine-5-)-Methyltransferase 1/metabolism , DNA Methylation/drug effects , Hepatic Stellate Cells/drug effects , Hepatic Stellate Cells/metabolism , Liver/drug effects , Liver/pathology , Liver Cirrhosis/chemically induced , Liver Cirrhosis/genetics , Liver Cirrhosis/metabolism , Macrophages/drug effects , Macrophages/metabolism , Male , Mice, Inbred C57BL , Rats , Sennosides/pharmacology , Up-Regulation/drug effectsABSTRACT
ETHNOPHARMACOLOGICAL RELEVANCE: Ershiwuwei Lvxue Pill (ELP, à½à½à¾²à½²à½à¼à½à½à½£à¼à½à½ºà½¢à¼à½£à¾à¼), a traditional Tibetan medicine preparation, has been used hundreds of years for the clinical treatment of rheumatoid arthritis (RA) in the highland region of Tibet, China. However, the underlying mechanism of its therapeutic effect remains unclear. AIM OF THE STUDY: The present study aimed to investigate the potential pharmacological mechanisms of anti-arthritic effect of ELP. MATERIALS AND METHODS: The main chemical constituents of ELP were analyzed by ultra-performance liquid chromatography quadrupole-time-flight mass spectrometry (UPLC-Q-TOF-MS). Forty-eight male Wistar rats (220 ± 20 g) were randomly divided into six groups: normal group, collagen-induced arthritis (CIA) group, methotrexate group (1.05 mg/kg), ELP groups (115, 230 and 460 mg/kg). CIA rat models were assigned to evaluate the anti-RA activity of ELP by determining the paws swelling, arthritis score, organ coefficients of spleen and thymus, and histopathological analysis of knee joints of synovial tissues. The levels of TNF-α, IL-10, IL-6 and IL-17 in serum were measured by ELISA. In addition, mRNA and protein expression levels associated with JAK2/STAT3 signaling pathway in synovial tissues of CIA rats were detected by qRT-PCR, immunohistochemistry and Western blot analyses. RESULTS: Fourteen main chemical constituents of ELP were quantitatively determined by UPLC-Q-TOF-MS analysis. Treatment with ELP reduced the paw swelling, arthritis score and organ coefficients of spleen and thymus. Histopathological examination revealed the protective effects of ELP on CIA rats with knee joint injury. The levels of serum pro-inflammatory cytokines (TNF-α, IL-6 and IL-17) were markedly reduced while the anti-inflammatory cytokine IL-10 was significantly increased with the treatment of ELP. Further investigations showed ELP down-regulated the mRNA and protein expression levels of Bcl-2, whereas up-regulated Bax, SOCS1 and SOCS3. Meanwhile, the ratios of p-JAK2/JAK2 and p-STAT3/STAT3 proteins from synovial tissues were dramatically decreased with the treatment of ELP, whereas no changes of the mRNA and protein expression levels of JAK2 and STAT3 were observed. CONCLUSION: These results indicated that ELP reduced the severity of arthritis and joint swelling, suggesting an antirheumatic effect on CIA rats. The possible mechanism is related to inhibiting inflammatory response and inducing apoptosis in synovial tissues by regulating JAK2/STAT3 signaling pathway. However, further in vivo and in vitro investigations are still needed to clarify the underlying mechanism of ELP in treating RA.
Subject(s)
Anti-Inflammatory Agents/pharmacology , Antirheumatic Agents/pharmacology , Arthritis, Experimental/drug therapy , Janus Kinase 2/antagonists & inhibitors , Medicine, Tibetan Traditional , Plant Extracts/pharmacology , STAT3 Transcription Factor/antagonists & inhibitors , Animals , Anti-Inflammatory Agents/chemistry , Anti-Inflammatory Agents/therapeutic use , Antirheumatic Agents/chemistry , Antirheumatic Agents/therapeutic use , Arthritis, Experimental/metabolism , Cytokines/metabolism , Janus Kinase 2/genetics , Janus Kinase 2/metabolism , Joints/pathology , Male , Plant Extracts/chemistry , Plant Extracts/therapeutic use , Proto-Oncogene Proteins c-bcl-2/genetics , Proto-Oncogene Proteins c-bcl-2/metabolism , Rats, Wistar , STAT3 Transcription Factor/genetics , STAT3 Transcription Factor/metabolism , Signal Transduction/drug effects , Suppressor of Cytokine Signaling 1 Protein/genetics , Suppressor of Cytokine Signaling 1 Protein/metabolism , Suppressor of Cytokine Signaling 3 Protein/genetics , Suppressor of Cytokine Signaling 3 Protein/metabolism , Synovial Membrane/drug effects , Synovial Membrane/metabolism , bcl-2-Associated X Protein/genetics , bcl-2-Associated X Protein/metabolismABSTRACT
BACKGROUND: Activation of renal fibroblasts is a critical mechanism in the process of renal fibrosis. As a commonly used herbal formula, Shenkang (SK) has been found to attenuate renal fibrosis and renal parenchyma destruction. However, the effect of SK on renal fibroblast activation in unilateral ureteral obstruction (UUO) mice and its molecular mechanism remain undetermined. The present study was performed to elucidate the effect of SK on renal fibroblast activation and renal fibrosis, as well as the potential underlying mechanism, in both NRK-49F cells and UUO mice. METHODS: NRK-49F cells were stimulated with 10 ng/ml TGF-ß1 for 48 h. After SK treatment, the CCK-8 method was used to evaluate cell viability. Thirty-six C57BL/6 mice were randomly divided into the sham group, UUO group, angiotensin receptor blocker (ARB) group, and SK high-, moderate- and low-dose groups. UUO was induced in mice except those in the sham group. Drugs were administered 1 day later. On the 13th day, the fractional anisotropy (FA) value was determined by MRI to evaluate the degree of renal fibrosis. After 14 days, serum indexes were assessed. Hematoxylin and eosin (HE) and Sirius red staining were used to observe pathological morphology and the degree of fibrosis of the affected kidney. Western blotting and PCR were used to assess the expression of related molecules in both cells and animals at the protein and gene levels. RESULTS: Our results showed that SK reduced extracellular matrix (ECM) and α-smooth muscle actin (α-SMA) expression both in vitro and in vivo and attenuated renal fibrosis and the pathological lesion degree after UUO, suppressing JAK2/STAT3 activation. Furthermore, we found that SK regulated the JAK2/STAT3 pathway regulators peroxiredoxin 5 (Prdx5) in vitro and suppressor of cytokine signaling protein 1 (SOCS1) and SOCS3 in vivo. CONCLUSIONS: These results indicated that SK inhibited fibroblast activation by regulating the JAK2/STAT3 pathway, which may be a mechanism underlying its protective action in renal fibrosis.
Subject(s)
Drugs, Chinese Herbal/therapeutic use , Fibroblasts/drug effects , Janus Kinase 2/metabolism , Nephrosclerosis/drug therapy , STAT3 Transcription Factor/metabolism , Actins/metabolism , Animals , Cell Line , Diffusion Tensor Imaging , Drug Evaluation, Preclinical , Drugs, Chinese Herbal/pharmacology , Extracellular Matrix/metabolism , Fibroblasts/metabolism , Fibrosis , Kidney/diagnostic imaging , Kidney/drug effects , Kidney/pathology , Male , Mice, Inbred C57BL , Nephrosclerosis/pathology , Peroxiredoxins/metabolism , Phytotherapy , Rats , Suppressor of Cytokine Signaling 1 Protein/metabolism , Suppressor of Cytokine Signaling 3 Protein/metabolism , Transforming Growth Factor beta1 , Ureteral ObstructionABSTRACT
ETHNOPHARMACOLOGICAL RELEVANCE: Pinellia pedatisecta Schott extract (PE) is generated from Pinellia pedatisecta Schott, a traditional Chinese medicinal plant. PE suppresses cervical tumor growth and exhibits effects on dendritic cells (DCs) that lead to modulation of antitumor CD4+ and CD8+ responses. AIMS: To explore the underlying mechanisms by which PE modulates tumor-associated dendritic cell (TADC) activation and function. METHODS: DCs and TADCs were generated from murine bone marrow and exposed to PE solutions at different doses, as well as to repeated doses separated at different time intervals. Quantitative PCR, Western blot analysis, flow cytometry, and gene silencing were used to analyze the modulatory effects of PE on the SOCS1/JAK2/STAT pathways. Furthermore, we separated human cervical tumor-infiltrated DCs (TIDCs) and conducted an ex-vivo stimulation model to observe the effect of PE. For phenotypic analysis of cultured DCs and ex vivo human specimens, we used flow cytometry to detect the molecular markers associated with cell function. RESULTS: In cultured TADCs and human cervical TIDCs, maturation- and functional markers (MHCII, CD80, CD83, CD86, and IL-12) were downregulated, whereas SOCS1 was upregulated. PE enhanced the expression of CD80, CD86, and IL-12 in cervical TIDCs, which induced increased expression of CD107a, GZMB, and perforin in CTLs, and furthermore induced apoptosis in a larger number of tumor cells. In cultured TADCs, PE downregulated SOCS1 expression and activated the phosphorylation of JAK2, STAT1, STAT4, and STAT5 in both dose- and time-dependent manners. The effects of PE upregulating MHCII, CD80, CD86, IL-12 on TADCs were blocked after SOCS1 silencing. CONCLUSIONS: In this study, PE restored the impaired function of cervical TIDCs, thereby eliciting further antitumor CTL responses. The effects of PE on TADCs were mediated through inhibition of SOCS1 and activation of downstream JAK2-STAT1/STAT4/STAT5 pathways. PE may be a potent and effective immunomodulatory drug for antitumor treatment via the blockade of SOCS1 signaling in DCs.
Subject(s)
Antineoplastic Agents, Phytogenic/pharmacology , Dendritic Cells/drug effects , Immunologic Factors/pharmacology , Pinellia , Plant Extracts/pharmacology , Suppressor of Cytokine Signaling 1 Protein/metabolism , Uterine Cervical Neoplasms/drug therapy , Animals , Antineoplastic Agents, Phytogenic/isolation & purification , Cell Line, Tumor , Dendritic Cells/immunology , Dendritic Cells/metabolism , Female , Gene Expression Regulation, Neoplastic , Humans , Immunologic Factors/isolation & purification , Janus Kinase 2/metabolism , Lipids/chemistry , Mice , Mice, Inbred C57BL , Pinellia/chemistry , Plant Extracts/isolation & purification , STAT Transcription Factors/metabolism , Signal Transduction , Solvents/chemistry , Suppressor of Cytokine Signaling 1 Protein/genetics , Tumor Cells, Cultured , Uterine Cervical Neoplasms/immunology , Uterine Cervical Neoplasms/metabolism , Uterine Cervical Neoplasms/pathologyABSTRACT
ETHNOPHARMACOLOGICAL RELEVANCE: Rhamnella gilgitica Mansf. et Melch. (སེà½à¼à½£à¾¡à½ºà½à¼à¼, RG) is a traditional Tibetan medicinal plant that is currently grown throughout Tibet. According to the theory of Tibetan medicine, RG is efficient for removing rheumatism, reducing swelling, and relieving pain. Hence, it has been used for the treatment of rheumatoid arthritis (RA) in Tibet for many years. However, there are no previous reports on the anti-RA activities of ethyl acetate extract of RG (RGEA). AIM OF THE STUDY: This study aimed to explore the anti-RA effect and mechanism of RGEA on collagen-induced arthritis (CIA) in rats. MATERIALS AND METHODS: The CIA model was established in male Wister rats by intradermal injection of bovine type II collagen and Complete Freund's Adjuvant at the base of the tail and left sole, respectively. The rats were orally administered with RGEA (9.71, 19.43, or 38.85 mg/kg) for 23 days. The body weight, swelling volume, arthritis index score, thymus and spleen indices, and pathological changes were observed to evaluate the effect of RGEA on RA. Furthermore, the inflammatory cytokines in serum, such as interleukin1 beta (IL-1ß), tumor necrosis factor alpha (TNF-α), interleukin6 (IL-6), interleukin17 (IL-17), interferon-γ (INF-γ), interleukin4 (IL-4), and interleukin10 (IL-10) were measured by enzyme linked immunosorbent assay (ELISA) to explore the anti-inflammatory effects of RGEA. The terminal deoxynucleotidyl transferase dUTP nick-end labelling (TUNEL) staining was used to examine apoptosis. Finally, the protein and gene expression of B-cell lymphoma-2-associated X (Bax), B-cell lymphoma 2 (Bcl-2), Caspase3, janus-activated kinase 2 (JAK2), signal transducer and activator of transcription 3 (STAT3), suppressor of cytokine signaling1 (SOCS1), and 3 (SOCS3) in synovial tissue were detected using immunohistochemistry and real-time quantitative polymerase chain reaction (RT-qPCR). RESULTS: After the treatment with RGEA, the body weight of rats was restored, both the arthritis index and paw swelling were suppressed, and spleen and thymus indices were decreased. RGEA reduced the inflammatory cells and synovial hyperplasia in the synovial tissue of the knee joint, and suppressed bone erosion. Meanwhile, RGEA decreased the levels of IL-1ß, IL-6, IL-17, TNF-α, and INF-γ, while increased the levels of IL-4 and IL-10. TUNEL fluorescence apoptosis results confirmed that RGEA obviously promoted the apoptosis of synovial cells. Further studies showed that RGEA inhibited the proteins and mRNAs expression of JAK2 and STAT3 as well as increased the proteins and mRNAs expression of SOCS1 and SOCS3. In addition, RGEA upregulated the expression of Bax and Caspase3, and downregulated the expression of Bcl-2. CONCLUSION: The anti-RA effectof RGEA might be related to the promotion of apoptosis and inhibition of inflammation, which regulated the JAK-STAT pathway.
Subject(s)
Antirheumatic Agents/pharmacology , Arthritis, Experimental/prevention & control , Janus Kinase 2/metabolism , Joints/drug effects , Plant Extracts/pharmacology , Rhamnaceae , STAT3 Transcription Factor/metabolism , Acetates/chemistry , Animals , Antirheumatic Agents/isolation & purification , Apoptosis/drug effects , Apoptosis Regulatory Proteins/genetics , Apoptosis Regulatory Proteins/metabolism , Arthritis, Experimental/chemically induced , Arthritis, Experimental/enzymology , Arthritis, Experimental/pathology , Collagen Type II , Cytokines/metabolism , Inflammation Mediators/metabolism , Janus Kinase 2/genetics , Joints/enzymology , Joints/pathology , Male , Medicine, Tibetan Traditional , Plant Extracts/isolation & purification , Rats, Wistar , Rhamnaceae/chemistry , STAT3 Transcription Factor/genetics , Signal Transduction , Solvents/chemistry , Suppressor of Cytokine Signaling 1 Protein/genetics , Suppressor of Cytokine Signaling 1 Protein/metabolism , Suppressor of Cytokine Signaling 3 Protein/genetics , Suppressor of Cytokine Signaling 3 Protein/metabolismABSTRACT
Inflammatory response that occur post-ischemia is a serious problem in the treatment of ischemic brain disease. MicroRNA-155 is a brain-specific or brain-enriched miRNA, which mediates inflammatory reactions in cerebral ischemic tissue by regulating inflammatory signal and the expression level of SOCS1. The present study was aimed to assess the effect of GuaLou GuiZhi Decoction (GLGZD) on miR-155 expression in activated microglia following inflammation and further explore the role of GLGZD on expression of the inflammation-related gene. BV2 cells were used to simulated by LPS to make the inflammatory model. Expression level of miR-155 was detected by Real-Time PCR. BV2 cells after simulated by LPS were then transfected with miR-155 mimic and its negative controls. Cytokines release were measured by corresponding purchased ELISA kits, respectively. Then target protein expression of miR-155 were detected by western blotting assay. After miRNA over expression transfections, expressions of inflammation-related factors, SOCS-1 and SAMD in BV2 cells after activation were measured by Western blot assay. Results showed that in BV2 cells after simulated by LPS, miR-155 was upregulated. The elevated miR-155 expression enhanced the inflammatory cytokine release. miR-155 directly target and negatively regulated SOCS-1 and SMAD-1 expression. Over expression of SOCS-1 and SMAD reduced inflammatory action that was enhanced by miR-155 mimic transfection. miR-155 was positively related with activation of NF-Ï°B signal pathways via SOCS-1 and SMAD. In conclusion, GuaLou GuiZhi Decoction (GLGZD) might exert its anti-inflammatory action by inhibiting the expression of miR-155, indicating that miR-155 may be used as a treatment target in clinical treatment with GuaLou GuiZhi Decoction (GLGZD) in ischemic brain.
Subject(s)
Anti-Inflammatory Agents/pharmacology , Drugs, Chinese Herbal/pharmacology , Lipopolysaccharides/pharmacology , MicroRNAs/physiology , Microglia/drug effects , Animals , Brain Ischemia/drug therapy , Cells, Cultured , Cytokines/biosynthesis , Mice , MicroRNAs/antagonists & inhibitors , Microglia/physiology , Smad1 Protein/antagonists & inhibitors , Suppressor of Cytokine Signaling 1 Protein/antagonists & inhibitorsABSTRACT
Triptolide( TP) is isolated from the traditional Chinese medicine Tripterygium wilfordii,which exhibits notable immuneregulative effect. Th17 cells involve in inflammatory response and Treg cells contribute to immune tolerance. They both play an important role in immune response. Previous studies have investigated that TP induced hepatic Th17/Treg imbalance. However,the effect of TP on spleen Th17/Treg cells remains unclear. Therefore,the aim of present study was to investigate the effect of TP on Th17/Treg cells in spleen. In this study,the effect of TP on the proliferation of splenic lymphocyte was detected by cytotoxicity test in vitro. After different concentrations of TP( 2. 5,5,20,40 nmol·L~(-1)) were given to splenic lymphocyte,cytokines secreted from the supernatant of splenic lymphocyte were detected by cytometric bead array,and the expression of suppressor of cytokine signaling( SOCS) mRNA was detected by qRT-PCR. Female C57 BL/6 mice were continuously observed for 24 h after treatment of 500 µg·kg-1 TP. The effects of TP on the splenic tissue structure and the percentage of Th17/Treg cells were examined. The results showed that the IC50 of TP was19. 6 nmol·L~(-1) in spleen lymphocytes. TP inhibited the secretion of IL-2 and IL-10 and induced the expression of SOCS-1/3 mRNA in spleen lymphocytes at the dosage of 2. 5 and 5 nmol·L~(-1) after 24 h in vitro. Administration of TP at dosage of 500 µg·kg-1 had no significant spleen toxicity in vivo. TP treatment increased the percentage of Th17 cells after 12 h and inhibited the proportion of Treg cells after 12 and 24 h. In conclusion,TP reduced the secretion of IL-2 and IL-10 through SOCS-1/3 signaling pathway,thereby induced the percentage of Th17 cells and inhibited the percentage of Treg cells.
Subject(s)
Diterpenes/pharmacology , Phenanthrenes/pharmacology , Spleen/drug effects , T-Lymphocytes, Regulatory/cytology , Th17 Cells/cytology , Animals , Cytokines/metabolism , Epoxy Compounds/pharmacology , Female , Mice , Mice, Inbred C57BL , Signal Transduction , Spleen/cytology , Suppressor of Cytokine Signaling 1 Protein/metabolism , Suppressor of Cytokine Signaling 3 Protein/metabolismABSTRACT
Triptolide( TP) is isolated from the traditional Chinese medicine Tripterygium wilfordii,which exhibits notable immuneregulative effect. Th17 cells involve in inflammatory response and Treg cells contribute to immune tolerance. They both play an important role in immune response. Previous studies have investigated that TP induced hepatic Th17/Treg imbalance. However,the effect of TP on spleen Th17/Treg cells remains unclear. Therefore,the aim of present study was to investigate the effect of TP on Th17/Treg cells in spleen. In this study,the effect of TP on the proliferation of splenic lymphocyte was detected by cytotoxicity test in vitro. After different concentrations of TP( 2. 5,5,20,40 nmol·L~(-1)) were given to splenic lymphocyte,cytokines secreted from the supernatant of splenic lymphocyte were detected by cytometric bead array,and the expression of suppressor of cytokine signaling( SOCS) mRNA was detected by qRT-PCR. Female C57 BL/6 mice were continuously observed for 24 h after treatment of 500 μg·kg-1 TP. The effects of TP on the splenic tissue structure and the percentage of Th17/Treg cells were examined. The results showed that the IC50 of TP was19. 6 nmol·L~(-1) in spleen lymphocytes. TP inhibited the secretion of IL-2 and IL-10 and induced the expression of SOCS-1/3 mRNA in spleen lymphocytes at the dosage of 2. 5 and 5 nmol·L~(-1) after 24 h in vitro. Administration of TP at dosage of 500 μg·kg-1 had no significant spleen toxicity in vivo. TP treatment increased the percentage of Th17 cells after 12 h and inhibited the proportion of Treg cells after 12 and 24 h. In conclusion,TP reduced the secretion of IL-2 and IL-10 through SOCS-1/3 signaling pathway,thereby induced the percentage of Th17 cells and inhibited the percentage of Treg cells.
Subject(s)
Animals , Female , Mice , Cytokines , Metabolism , Diterpenes , Pharmacology , Epoxy Compounds , Pharmacology , Mice, Inbred C57BL , Phenanthrenes , Pharmacology , Signal Transduction , Spleen , Cell Biology , Suppressor of Cytokine Signaling 1 Protein , Metabolism , Suppressor of Cytokine Signaling 3 Protein , Metabolism , T-Lymphocytes, Regulatory , Cell Biology , Th17 Cells , Cell BiologyABSTRACT
Berberine has been reported to have protective effects in colitis treatment. However, the detailed mechanisms remain unclear. Herein, we demonstrated that berberine could protect against dextran sulfate sodium (DSS)-induced colitis in mice by regulating macrophage polarization. In the colitis mouse model, berberine ameliorated DSS-induced colon shortening and colon tissue injury. Moreover, berberine-treated mice showed significant reduction in the disease activity index (DAI), pro-inflammatory cytokines expression and macrophages infiltration compared with the DSS-treated mice. Notably, berberine significantly reduced the percentage of M1 macrophages. In vitro analysis also confirmed the inhibitory effects of berberine on macrophages M1 polarization in RAW267.4 cells. Further investigation showed that berberine promoted AKT1 expression in mRNA and protein level. Silence of AKT1 abolished the inhibitory effect of berberine on macrophages M1 polarization. The berberine-induced AKT1 expression promoted suppressers of cytokine signaling (SOCS1) activation, which inhibited nuclear factor-kappa B (NF-κB) phosphorylation. In addition, we also found that berberine activated AKT1/SOCS1 signaling pathway but inhibited p65 phosphorylation in macrophages in vivo. Therefore, we concluded that berberine played a regulatory role in macrophages M1 polarization in DSS-induced colitis via AKT1/SOCS1/NF-κB signaling pathway. This unexpected property of berberine may provide a potential explanation for its protective effects in colitis treatment.
Subject(s)
Anti-Inflammatory Agents/therapeutic use , Berberine/therapeutic use , Cell Differentiation/drug effects , Colitis/drug therapy , Macrophages/physiology , Animals , Colitis/chemically induced , Cytokines/metabolism , Dextran Sulfate , Disease Models, Animal , Humans , Inflammation Mediators/metabolism , Male , Mice , Mice, Inbred C57BL , NF-kappa B/metabolism , Proto-Oncogene Proteins c-akt/genetics , Proto-Oncogene Proteins c-akt/metabolism , RAW 264.7 Cells , Signal Transduction , Suppressor of Cytokine Signaling 1 Protein/genetics , Suppressor of Cytokine Signaling 1 Protein/metabolism , Th1 Cells/immunologyABSTRACT
Decreased interferon (IFN)-γ levels and increased levels of macrophage-derived chemokine (MDC) and intercellular adhesion molecule (ICAM)-1 are known to be involved in allergic skin diseases, such as eczema and atopic dermatitis. Activation of the IFN-γ and its downstream interleukin-12 (IL-12) pathway can correct these diseases. Suppressor of cytokine signaling 1 (SOCS1) is a cytokine signaling inhibitor that blocks downstream pathways of IFN-γ by blocking the mitogen-activated protein kinase (MAPK) and protein kinase B (Akt) signaling pathways. Oxymatrine (OMT), a quinolizidine alkaloid extracted from the herbal medicine Radix Sophorae flavescentis, is used to treat allergic skin diseases in China. The non-cytotoxic concentrations of OMT in HaCaT cells were determined through 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assays. Tumor necrosis factor (TNF)-α and IFN-γ were used to stimulate HaCaT cells, and OMT was added to this system with tacrolimus (FK506) as a positive control. The mRNAs of cytokines, MDC, ICAM-1, IL-12p35, IL-12p40, and IFN-γ receptor (IFN-γR)α were detected by RT-PCR. Western blot analyses were performed to assess activation of the MAPK (p38, Jun N-terminal kinase, and extracellular signal-regulated kinase) and Akt signaling pathways. OMT increased the mRNA levels of the IL-12 and IFN-γRα, reduced the mRNA levels of ICAM-1, MDC, and SOCS1. But FK506 increased the mRNA levels of IL12 and inhibited the expression of ICAM-1 mRNAs and had no effects on the IFN-γRα, MDC, and SOCS1 mRNA in HaCaT cells stimulated with TNF-α and IFN-γ. Thus, the mechanisms through which OMT and FK506 ameliorate allergic skin diseases differ.
Subject(s)
Alkaloids/pharmacology , Quinolizines/pharmacology , Skin Diseases/drug therapy , Cell Line , Chemokine CCL22 , Down-Regulation/drug effects , Humans , Immunosuppressive Agents , Intercellular Adhesion Molecule-1 , Interferon-gamma/metabolism , Keratinocytes/cytology , MAP Kinase Signaling System , RNA, Messenger/drug effects , Skin Diseases/immunology , Suppressor of Cytokine Signaling 1 Protein , Tacrolimus/pharmacologyABSTRACT
Macrophage polarization is flexible, and involves in different signaling pathways and various transcription factors. Suppressor of cytokine signaling (SOCS) is an important inhibitor of cytokine signaling pathways and also a key physiological regulator for natural and acquired immunity systems. Following transfection of SOCS1 short hairpin (sh)RNA into mouse macrophage cells, reverse transcriptionquantitative polymerase chain reaction demonstrated that the mRNA levels of Janus kinase (JAK)1 and signal transducer and activator of transcription (STAT)1 increased significantly. In addition, western blotting indicated that JAK1, STAT1 and pSTAT1 expression was significantly enhanced. Fludarabine can inhibit phosphorylation of STAT1 and SOCS1 expression. When fludarabine was added and SOCS1 shRNA was transfected, the inhibition of fludarabine was weakened, and pSTAT1 expression was upregulated. Flow cytometry detection indicated that, following the downregulation of SOCS1 expression, M1type cells significantly increased, but the proportion of M2type cells did not change significantly. Fludarabine can reduce the effect of SOCS1 shRNA on promoting M1type cell polarization, and macrophages can polarize into both M1 and M2 phenotypes. Further ELISA results presented that, when downregulating SOCS1 expression, interleukin (IL)4 and IL10 expression was both downregulated, and tumor necrosis factor (TNF)α and interferon (IFN)γ expression was significantly upregulated. When adding fludarabine or injecting with the traditional Chinese medicine Xuebijing, IL4 and IL10 expression was both significantly upregulated, and TNFα and IFNγ expression was significantly downregulated. When adding fludarabine and downregulating SOCS1, IL4, IL10, TNFα and IFNγ expression presented no significant changes. The above results indicated that, when SOCS1 expression is downregulated, it will activate the JAK1/STAT1 pathway, and thereby promote the polarization of macrophages into M1 type. The findings are of great importance for understanding occurrence, development and treatment of various immunerelated diseases.
Subject(s)
Janus Kinase 1/immunology , Macrophages, Peritoneal/immunology , STAT1 Transcription Factor/immunology , Signal Transduction/immunology , Suppressor of Cytokine Signaling 1 Protein/immunology , Animals , Anti-Inflammatory Agents/pharmacology , Cell Differentiation , Drugs, Chinese Herbal/pharmacology , Enzyme Inhibitors/pharmacology , Gene Expression Regulation , Interferon-gamma/genetics , Interferon-gamma/immunology , Interleukin-10/genetics , Interleukin-10/immunology , Interleukin-4/genetics , Interleukin-4/immunology , Janus Kinase 1/genetics , Macrophages, Peritoneal/cytology , Macrophages, Peritoneal/drug effects , Mice , Phosphorylation , Primary Cell Culture , RNA, Small Interfering/genetics , RNA, Small Interfering/immunology , STAT1 Transcription Factor/agonists , STAT1 Transcription Factor/genetics , Suppressor of Cytokine Signaling 1 Protein/antagonists & inhibitors , Suppressor of Cytokine Signaling 1 Protein/genetics , Tumor Necrosis Factor-alpha/genetics , Tumor Necrosis Factor-alpha/immunology , Vidarabine/analogs & derivatives , Vidarabine/antagonists & inhibitors , Vidarabine/pharmacologyABSTRACT
BACKGROUND: The CpG island methylator phenotype (CIMP) in stage III colon cancer (CRC) has been associated with improved survival after treatment with adjuvant irinotecan-based chemotherapy. In this analysis, we determine whether CIMP status in the primary CRC is concordant with the CIMP status of matched metastases in order to determine if assessment of CIMP status in the primary tumor can be used to predict CIMP status of metastatic disease, which is relevant for patient management as well as for understanding the biology of CIMP CRCs. METHODS: We assessed the CIMP status of 70 pairs of primary CRC and matched metastases using a CRC-specific panel of five markers (CACNA1G, IGF2, NEUROG1, RUNX3, and SOCS1) where CIMP positive was defined as 3/5 positive markers at a percent methylated reference threshold of ≥10%. Concordance was compared using the Fisher's exact test and P < 0.05 was considered significant. RESULTS: Sixty-nine of the pairs (98.6%) showed concordant CIMP status in the primary tumor and matched metastasis; five (7.0%) of the pairs were concordantly CIMP positive. Only one pair (1.4%) had divergent CIMP status, demonstrating CIMP positivity (4/5 markers positive) in the primary tumor, while the matched metastasis was CIMP negative (0 markers positive). CONCLUSIONS: CIMP status is generally concordant between primary CRCs and matched metastases. Thus, CIMP status in the primary tumor is maintained in matched metastases and can be used to inform CIMP-based therapy options for the metastases.
Subject(s)
Biomarkers, Tumor/genetics , Colorectal Neoplasms/genetics , DNA Methylation , Adult , Aged , Basic Helix-Loop-Helix Transcription Factors/genetics , Calcium Channels, T-Type/genetics , Colorectal Neoplasms/pathology , Core Binding Factor Alpha 3 Subunit/genetics , CpG Islands , Epigenesis, Genetic , Female , Humans , Insulin-Like Growth Factor II/genetics , Male , Middle Aged , Neoplasm Metastasis , Nerve Tissue Proteins/genetics , Phenotype , Suppressor of Cytokine Signaling 1 Protein/geneticsABSTRACT
The immune system is critical in preventing infection and cancer, and malnutrition can weaken different aspects of the immune system to undermine immunity. Previous studies suggested that vitamin B6 deficiency could decrease serum antibody production with concomitant increase in IL4 expression. However, evidence on whether vitamin B6 deficiency would impair immune cell differentiation, cytokines secretion, and signal molecule expression involved in JAK/STAT signaling pathway to regulate immune response remains largely unknown. The aim of this study is to investigate the effects of vitamin B6 deficiency on the immune system through analysis of T lymphocyte differentiation, IL-2, IL-4, and INF-γ secretion, and SOCS-1 and T-bet gene transcription. We generated a vitamin B6-deficient mouse model via vitamin B6-depletion diet. The results showed that vitamin B6 deficiency retards growth, inhibits lymphocyte proliferation, and interferes with its differentiation. After ConA stimulation, vitamin B6 deficiency led to decrease in IL-2 and increase in IL-4 but had no influence on IFN-γ. Real-time PCR analysis showed that vitamin B6 deficiency downregulated T-bet and upregulated SOCS-1 transcription. This study suggested that vitamin B6 deficiency influenced the immunity in organisms. Meanwhile, the appropriate supplement of vitamin B6 could benefit immunity of the organism.
Subject(s)
Cytokines/genetics , T-Lymphocytes/immunology , T-Lymphocytes/physiology , Vitamin B 6 Deficiency/immunology , Animals , Cell Differentiation , Diet , Down-Regulation , Interferon-gamma/genetics , Interferon-gamma/metabolism , Interleukin-2/genetics , Interleukin-2/metabolism , Interleukin-4/genetics , Interleukin-4/metabolism , Lymphocyte Activation , Mice , Pyridoxal Phosphate/analogs & derivatives , Pyridoxal Phosphate/blood , Suppressor of Cytokine Signaling 1 Protein/genetics , T-Box Domain Proteins/genetics , Vitamin B 6 Deficiency/blood , Vitamin B 6 Deficiency/metabolism , Xanthurenates/bloodABSTRACT
Streptococcus pneumoniae is a major human pathogen and a leading cause of pneumonia, septicemia, and meningitis worldwide. Despite clinical studies linking vitamin D deficiency and pneumonia, molecular mechanisms behind these observations remain unclear. In particular, the effects of vitamin D on neutrophil responses remain unknown. Using pneumococcal strains, primary neutrophils isolated from human blood, and sera from patients with frequent respiratory tract infections (RTIs), we investigated the effects of vitamin D on neutrophil bactericidal and inflammatory responses, including pattern recognition receptors, antimicrobial peptides, and cytokine regulation. We found that vitamin D upregulated pattern recognition receptors, TLR2, and NOD2, and induced the antimicrobial human neutrophil peptides (HNP1-3) and LL-37, resulting in increased killing of pneumococci in a vitamin D receptor-dependent manner. Antibodies targeting HNP1-3 inhibited bacterial killing. Vitamin D supplementation of serum from patients with bacterial RTIs enhanced neutrophil killing. Moreover, vitamin D lowered inflammatory cytokine production by infected neutrophils via IL-4 production and the induction of suppressor of cytokine signaling (SOCS) proteins SOCS-1 and SOCS-3, leading to the suppression of NF-κB signaling. Thus, vitamin D enhances neutrophil killing of S. pneumoniae while dampening excessive inflammatory responses and apoptosis, suggesting that vitamin D could be used alongside antibiotics when treating pneumococcal infections.
Subject(s)
Inflammation/drug therapy , Neutrophils/immunology , Pneumococcal Infections/drug therapy , Streptococcus pneumoniae/immunology , Vitamin D/pharmacology , Bacteriolysis , Cells, Cultured , Humans , Immunomodulation , Interleukin-4/metabolism , NF-kappa B/metabolism , Nod2 Signaling Adaptor Protein/genetics , Nod2 Signaling Adaptor Protein/metabolism , Primary Cell Culture , Signal Transduction , Suppressor of Cytokine Signaling 1 Protein/genetics , Suppressor of Cytokine Signaling 1 Protein/metabolism , Suppressor of Cytokine Signaling Proteins/genetics , Suppressor of Cytokine Signaling Proteins/metabolism , Toll-Like Receptor 2/genetics , Toll-Like Receptor 2/metabolism , alpha-Defensins/genetics , alpha-Defensins/metabolismABSTRACT
Objective To explore the relationship between Xinfeng Capsule (XFC) improving the hypercoagulative state in patients with Sjogren's syndrome (SS) and miR-155/suppressor of cytokine signaling 1 (SOCS1)/nuclear factor κB (NF-κB) signaling pathway. Methods Sixty-six SS patients were randomly divided into XFC-treated group and hydroxychloroquine (HCQ)-treated control group (n=33 per group), which were respectively treated with XFC and HCQ. In addition, 20 healthy volunteers were enrolled as a normal control group. The levels of prothrombin time (PT), activated partial thromboplastin time (APTT), fibrinogen (FIG), thrombin time (TT) and D-dimer (D-D) were detected using automatic coagulation analyzer. Interleukin-1ß (IL-1ß), IL-4, IL-10, tumor necrosis factor-α (TNF-α), P50, P65, inhibitor of NF-κB α (IκBα) were tested using ELISA. Meanwhile, the mRNA expressions of p50, p65 and IκBα were determined using quantitative real-time PCR, and the level of microRNA-155 (miR-155) was examined by one-step fluorescence quantitative PCR. The protein levels of P50, P65 and SOCS1 were detected using Western blotting. Erythrocyte sedimentation rate (ESR) was evaluated by Westergren method. Hypersensitive C-reactive protein (hs-CRP) was detected using automatic biochemical analyzer. Results Compared with the normal control group, the levels of D-D and FIB significantly increased in SS group; simultaneously, the serum levels of miR-155, IL-1ß, TNF-α, P50, P65, IκBα, hs-CRP, ESR were significantly elevated in SS patients, while IL-4 and IL-10 were significantly reduced. Spearman correlation analysis showed that the coagulation parameters were remarkably correlated with cytokines, NF-κB and activity indexes. In the two treated groups, coagulation parameters and related indexes were demonstrated having some improvement, especially in the XFC group, which had a much higher efficiency, and better outcomes in reducing the levels of FIB, D-D, miR-155, TNF-α, IL-1ß, P50, P65, ESR and hs-CRP, as well as increasing the expressions of SOCS1, IL-4 and IL-10. Conclusion XFC can significantly alleviate the hypercoagulative state of patients with SS, and the mechanisms may be related to the inhibition of miR-155/SOCS1/NF-κB signaling pathway.
Subject(s)
Blood Coagulation/drug effects , Drugs, Chinese Herbal/administration & dosage , MicroRNAs/metabolism , NF-kappa B/metabolism , Sjogren's Syndrome/drug therapy , Sjogren's Syndrome/metabolism , Suppressor of Cytokine Signaling 1 Protein/metabolism , Adult , Aged , Female , Humans , Interleukin-10/genetics , Interleukin-10/metabolism , Interleukin-4/genetics , Interleukin-4/metabolism , Male , MicroRNAs/genetics , Middle Aged , NF-kappa B/genetics , Signal Transduction/drug effects , Sjogren's Syndrome/blood , Sjogren's Syndrome/genetics , Suppressor of Cytokine Signaling 1 Protein/geneticsABSTRACT
Reducing ß amyloid- (Aß-) induced microglial activation is believed to be effective in treating Alzheimer's disease (AD). Microglia can be activated into classic activated state (M1 state) or alternative activated state (M2 state), and the former is harmful; in contrast, the latter is beneficial. Gypenoside (GP) is the major bioactive constituent of Gynostemma pentaphyllum, a traditional Chinese herb medicine. In this study, we hypothesized that GP attenuates Aß-induced microglial activation by ameliorating microglial M1/M2 states, and the process may be mediated by suppressor of cell signaling protein 1 (SOCS1). In this study, we found that Aß exposure increased the levels of microglial M1 markers, including iNOS expression, tumor necrosis factor α (TNF-α), interleukin 1ß (IL-1ß), and IL-6 releases, and coadministration of GP reversed the increase of M1 markers and enhanced the levels of M2 markers, including arginase-1 (Arg-1) expression, IL-10, brain-derived neurotrophic factor (BDNF), and glial cell-derived neurotrophic factor (GDNF) releases in the Aß-treated microglial cells. SOCS1-siRNA, however, significantly abolished the GP-induced effects on the levels of microglial M1 and M2 markers. These findings indicated that GP attenuates Aß-induced microglial activation by ameliorating M1/M2 states, and the process may be mediated by SOCS1.
Subject(s)
Amyloid beta-Peptides/pharmacology , Inflammation/metabolism , Microglia/drug effects , Signal Transduction/drug effects , Suppressor of Cytokine Signaling 1 Protein/metabolism , Animals , Brain-Derived Neurotrophic Factor/metabolism , Cell Line , Gynostemma , Inflammation/drug therapy , Interleukin-10/metabolism , Interleukin-1beta/metabolism , Interleukin-6/metabolism , Mice , Microglia/metabolism , Nitric Oxide Synthase Type I/metabolism , Plant Extracts/pharmacology , Plant Extracts/therapeutic use , Tumor Necrosis Factor-alpha/metabolismABSTRACT
BACKGROUND: Signal transducer and activator of transcription 3 (STAT3) is persistently activated in squamous cell carcinoma of the head and neck (SCCHN) and can cause uncontrolled cellular proliferation and division. HYPOTHESIS: Thus, its targeted abrogation could be an effective strategy to reduce the risk of SCCHN. Resveratrol is known for its anti-cancer efficacy in a variety of cancer models. STUDY DESIGN: The effect resveratrol on STAT3 activation, associated protein kinases, phosphatases, cellular proliferation and apoptosis was investigated. METHODS: We evaluated the effect of resveratrol on STAT3 signaling cascade and its regulated functional responses in SCCHN cells. RESULTS: We found that HN3 and FaDu cells expressed strongly phosphorylated STAT3 on both tyrosine 705 and serine 727 residues as compared to other SCCHN cells. The phosphorylation was completely suppressed by resveratrol in FaDu cells, but not substantially in HN3 cells. STAT3 suppression was mediated through the inhibition of activation of upstream JAK2, but not of JAK1 and Src kinases. Treatment with the protein tyrosine phosphatase (PTP) inhibitor pervanadate reversed the resveratrol-induced down-regulation of STAT3, thereby indicating a critical role for a PTP. We also found that resveratrol induced the expression of the SOCS-1 protein and mRNA. Further, deletion of SOCS-1 gene by siRNA suppressed the induction of SOCS-1, and reversed the inhibition of STAT3 activation. Resveratrol down-regulated various STAT3-regulated gene products, inhibited proliferation, invasion, as well as induced the cell accumulation in the sub-G1 phase and caused apoptosis. Beside, this phytoalexin also exhibited the enhancement of apoptosis when combined with ionizing radiation treatment. CONCLUSION: Our results suggest that resveratrol blocks STAT3 signaling pathway through induction of SOCS-1, thus attenuating STAT3 phosphorylation and proliferation in SCCHN cells.